Abstract Morbidity and mortality associated with Chagas disease in Latin America exceed better-known conditions such as malaria, tuberculosis, or AIDS. Millions of people are affected by this trypanosomiasis. No vaccines are available to prevent this disease and drug treatments have serious side effects and are not completely effective. Survival of Trypanosoma cruzi in the mammalian hosts depends on the parasite's ability to infect host cells, reproduce, and live in the blood of the host long enough to warrant its transmission through a bloodsucking insect vector. Our aim is to investigate these survival mechanisms and their role in the pathogenesis of T. cruzi infection, and shed light into potential ways to control the disease. Survival of T. cruzi trypomastigotes in the blood of the mammalian host and in the intestine of the vector depends in great part on their ability to tolerate dramatic changes in osmolarity during their circulation through the kidney medulla of the mammalian host (1,300-1,400 mOsm/Kg) or their passage through the rectum of the insect vector (1,000 mOsm/Kg), and we have found that polyphosphate (polyP) and the contractile vacuole complex (CVC) have en essential role in their survival mechanisms. Recent results from our laboratory suggest that polyP and/or pyrophosphate (PPi) could also be involved in the pathogenesis of T. cruzi infection. To reduce polyP levels we overexpressed a degradative enzyme that hydrolyzes both PPi and polyP resulting in a dramatic decrease of PPi/polyP. Mutant trypomastigotes overexpressing the enzyme did not produce detectable parasitemias and in several experiments all infected animals survived an otherwise lethal infection. Mice infected with parasites deficient in PPi/polyP failed to develop parasitemia after immunosuppression with cyclophosphamide strongly suggesting that infection had been completely cleared. Our results underscore an important role for PPi/polyP in the pathogenesis of T. cruzi infection. PolyP has been shown to act as a virulence factor in bacteria but little is known on its role in eukaryotic pathogens, besides its pro-coagulant and pro-inflammatory activities. Survival within the mammalian host also depends on the ability of T. cruzi to invade different host cells, escape from the parasitophorus vacuole and replicate intracellularly. T. cruzi trans-sialidases have been demonstrated to have essential roles in these mechanisms. However, the main obstacle in assessing the function of trans-sialidases is that knockout parasites were never obtained due to the large number of gene copies scattered through the genome. We have found that trans-sialidases traffic through the contractile vacuole in their way to the plasma membrane, and that disruption of this traffic by interfering with their passage through the CVC results in parasites devoid of these proteins in their surface. The study of this trafficking mechanism and of ways to interfere with this traffic will contribute to the understanding of the pathogenesis of T. cruzi infection.